Versatile broad-host-range cosmids for construction of high quality metagenomic libraries

Jiujun Cheng; Lee Pinnell; Katja Engel; Josh D Neufeld; Trevor C Charles

doi:10.1016/j.mimet.2014.01.015

Versatile broad-host-range cosmids for construction of high quality metagenomic libraries

J Microbiol Methods. 2014 Apr:99:27-34. doi: 10.1016/j.mimet.2014.01.015. Epub 2014 Feb 1.

Authors

Jiujun Cheng¹, Lee Pinnell¹, Katja Engel¹, Josh D Neufeld¹, Trevor C Charles²

Affiliations

¹ Department of Biology, University of Waterloo, 200 University Avenue West, Waterloo, Ontario N2L 3G1, Canada.
² Department of Biology, University of Waterloo, 200 University Avenue West, Waterloo, Ontario N2L 3G1, Canada. Electronic address: trevor.charles@uwaterloo.ca.

PMID: 24495694
DOI: 10.1016/j.mimet.2014.01.015

Abstract

We constructed IncP broad-host-range Gateway® entry cosmids pJC8 and pJC24, which replicate in diverse Proteobacteria. We demonstrate the functionality of these vectors by extracting, purifying, and size-selecting metagenomic DNA from agricultural corn and wheat soils, followed by cloning into pJC8. Metagenomic DNA libraries of 8×10(4) (corn soil) and 9×10(6) (wheat soil) clones were generated for functional screening. The DNA cloned in these libraries can be transferred from these recombinant cosmids to Gateway® destination vectors for specialized screening purposes. Those library clones are available from the Canadian MetaMicroBiome Library project (http://www.cm2bl.org/).

Keywords: Broad-host-range cosmids; DNA extraction; Functional metagenomics; Gateway® recombination vector; Metagenomic libraries; Soil.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cosmids*
Gene Library*
Genetics, Microbial / methods*
Host Specificity
Metagenomics / methods*
Molecular Biology / methods*
Proteobacteria / genetics*